CD8 T cells play crucial roles in control of infections with viruses, intracellular bacteria and protozoa. Acute viral infections typically induce a potent self-renewing population of memory CD8 T cells that persists for decades after infection. Durable protective immunity to repeated infectious insults depends upon the maintenance of a stem-cell-like self-renewing population of memory CD8 T cells with high regenerative capacity i.e. the competence to endure repeated cycles of clonal expansion and differentiation into effector and memory CD8 T cells without attaining senescence. However, human aging is associated with the progressive accumulation of senescent and terminally differentiated memory CD8 T cells (effector memory CD45RA+ve; EMRA) that display poor effector functions and regenerative capacity. The mechanisms that regulate the senescence and regenerative potential of memory CD8 T cells remain poorly understood. The FoxO family of transcription factors mitigates senescence and promotes the regenerative capacity of stem cells. By contrast, the tumor suppressor molecule p16INK4a promotes senescence and limits the regenerative potential of stem cells. Further, p16INK4a expression in T cells is a biomarker of aging, and proliferative arrest of terminally differentiated human T cells is linked to elevated p16INK4a levels. Exciting preliminary data show that: (1) FoxO1 expression is markedly reduced in human senescent EMRA T cells; (2) FoxO1 deficiency results in senescence, regenerative decline of memory CD8 T cells and elevated expression of p16INK4a in mice. Based on these data, in specific aim 1, we will test the hypothesis that FoxO1 opposes senescence and promotes the regenerative potential of memory CD8 T cells by repressing p16INK4a expression. Not only FoxO1 deficiency, aberrant activation of the mechanistic target of rapamycin (mTOR) is also a prominent feature of cellular senescence and aging. Remarkably, several activities of FoxOs faithfully phenocopy the effects of mTOR inhibition in T cells and stem cells, which supports the idea that the functions of FoxO also include antagonism of mTOR. Based on preliminary data that FoxO1 ablation elevated mTOR activity in memory CD8 T cells, in specific aim 2, we will test the hypothesis that FoxO1 opposes senescence and promotes the protective capacity of memory CD8 T cells by restraining mTOR activity. This exploratory R21 proposal is innovative because, it seeks to identify a new link between FoxO1, p16 and mTOR in governing the regenerative potential of memory CD8 T cells, and provide mechanistic insights into the dysregulated state and regenerative decline of memory T cells in the elderly. Such new insights into the tractable molecular pathways are predicted to have a substantive and broad impact on human health because, we envision development of strategies to therapeutically modulate the FoxO1-p16/mTOR axis to: (1) restore T-cell homeostasis in the elderly; (2) enhance vaccine-induced CD8 T cell memory; (3) elicit and maintain durable protective cell-mediated immunity in the elderly.